注射用糜蛋白酶溶解后稳定性及雾化粒径考察

Stability and Atomized Particle Size of Chymotrypsin for Injection After Dissolution

目的建立测定注射用糜蛋白酶溶解后含量的反相高效液相色谱(RP-HPLC)法,并考察注射用糜蛋白酶雾化吸入溶液(简称雾化溶液)的稳定性和雾化粒径分布。方法色谱柱为Grace 214TPC_(4)柱(250 mm×4.6 mm,5μm),流动相为0.1%三氟乙酸水溶液-0.09%三氟乙酸乙腈(梯度洗脱),流速为1.0 mL/min,检测波长为214 nm,柱温为30℃,进样量为10μL;采用RP-HPLC法测定雾化溶液0,0.5,1.0,2.0,4.0,6.0 h时的含量,并考察稳定性;采用SpraytecSTP2000型激光粒度仪测定雾化溶液的粒径。结果糜蛋白酶质量浓度在0.20~2.00 mg/mL范围内与峰面积线性关系良好,精密度、稳定性、重复性试验的RSD均不超过2.0%(n=5),平均回收率为(100.81±2.47)%,RSD为2.45%(n=9);注射用糜蛋白酶分别用灭菌注射用水和0.9%氯化钠注射液配制后,在4℃下6 h内均呈澄清状态,2 h内含量均较稳定,2,4,6 h内的平均降解率分别为(9.05±4.59)%和(13.51±3.99)%,(14.25±4.89)%和(15.45±3.19)%,(15.18±3.86)%和(14.08±4.84)%,均显著高于0.5 h时(P<0.05),且用灭菌注射用水配制雾化溶液的pH接近中性;用0.9%氯化钠注射液配制的雾化溶液,累积分布达50%时的粒径为(3.59±0.14)μm,显著低于用灭菌注射用水配制的粒径(P<0.05)。结论注射用糜蛋白酶雾化溶液的含量在接近中性条件下2h内较稳定,在0.9%氯化钠注射液及灭菌注射用水中雾化效果均可达到预期,且均到达肺部。

Objective To establish an reversed phase-high performance liquid chromatography(RP-HPLC)for the content determination of Chymotrypsin for Injection after dissolution,and to investigate the stability and the distribution of particle size of Chymotrypsin for Injection aerosol inhalation solution(hereinafter referred to as aerosol inhalation solution)after atomization.Methods The chromatographic column was Grace 214TP C4 column(250 mm×4.6 mm,5μm),the mobile phase was 0.1%acetic acid solution-0.09%trifluoroacetic acid acetonitrile(gradient elution),the flow rate was 1.0 mL/min,the detection wavelength was 214 nm,the column temperature was 30℃,and the injection volume was 10μL.The content of aerosol inhalation solution at 0,0.5,1.0,2.0,4.0 and 6.0 h was determined by the RP-HPLC method.The particle size of the aerosol inhalation solution was determined by Spraytec STP2000 laser particle sizer.Results The linear range of chymotrypsin was 0.20-2.00 mg/mL,The RSDs of precision,stability and repeatability tests were not move than 2.0%(n=5).The average recovery of chymotrypsin was(100.81±2.47)%with an RSD of 2.45%(n=9).After Chymotrypsin for Injection was prepared with Water for Sterilization Injection and 0.9%Sodium Chloride Injection respectively,all the aerosol inhalation solution was clear within 6 h at 4℃,the content was stable within 2 h,the average degradation rates of the aerosol inhalation solution prepared with two different solvents within 2,4 and 6 h were(9.05±4.59)%and(13.51±3.99)%,(14.25±4.89)%and(15.45±3.19)%,(15.18±3.86)%and(14.08±4.84)%respectively,which were significantly higher than those within 0.5 h(P<0.05),and the pH of the aerosol inhalation solution prepared with Water for Sterilization Injection was close to neutral solution.The DV(50)of Chymotrypsin for Injection dissolved and atomized with 0.9%Sodium Chloride Injection was(3.59±0.14)μm,which was significantly lower than that dissolved and atomized with Sterilized Water For Injection(P<0.05).Conclusion The content of Chymotrypsin for Injection aerosol inhalation solution is more stable in neutral condition within 2 h,and can be well atomized in 0.9%Sodium Chloride Injection and Water for Sterilization Injection to reach the lung.